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利用磁珠富集法和5’锚定PCR法开发背瘤丽蚌的微卫星标记,将获得的多态性引物用于群体的遗传多态性分析,以期在比较两种开发微卫星标记方法的基础上同时获得一批有用的微卫星引物。从磁珠富集法获得的微卫星序列阳性克隆率为69.2%,重复次数超过10的占总数的70.2%,从设计的28对引物中筛选得到多态性引物11对,开发效率为39.3%。这11对引物用于养殖群体的遗传多样性分析,结果显示,等位基因数范围为4～13,观测杂合度、期望杂合度范围分别为0.205～0.738、0.566～0.839。而5’锚定PCR法获得的微卫星序列阳性克隆率为97.8%,重复次数超过10的占总数的24.7%,从设计的56对引物中筛选得到多态性引物19对,开发效率为30.4%。这19对引物用于养殖群体的遗传多样性分析,结果显示,等位基因数范围为3～10,观测杂合度、期望杂合度范围分别为0.208～0.894、0.431～0.896。实验结果表明,磁珠富集法所获微卫星序列质量高,开发微卫星标记效率较高;而5’锚定PCR法实验操作更简便,所获得的引物遗传多样性指数更高。两种方法开发的引物均可用于背瘤丽蚌和近缘种的野生种质资源遗传多样性研究。
The microsatellite markers of Rehmannia polygamis were developed by magnetic bead enrichment and 5 ’anchored PCR, and the obtained polymorphic primers were used to analyze the genetic polymorphism of the population. In order to compare two microsatellite markers Based on the same time access to a number of useful microsatellite primers. Microsatellite sequences obtained by magnetic bead enrichment method was 69.2%, 70.2% of the total number of repeats, and 11 pairs of polymorphic primers were screened from 28 pairs of primers. The development efficiency was 39.3% . The 11 pairs of primers were used to analyze the genetic diversity of farmed populations. The results showed that the number of alleles ranged from 4 to 13, and the observed heterozygosity ranged from 0.205 to 0.738 and from 0.566 to 0.839, respectively. The positive rate of microsatellite loci obtained by 5 ’anchored PCR was 97.8%, and the number of repeats exceeded 10 accounted for 24.7% of the total. Nineteen pairs of primers were screened from 19 pairs of primers, and the development efficiency was 30.4 %. The 19 pairs of primers were used to analyze the genetic diversity of farmed populations. The results showed that the number of alleles ranged from 3 to 10 and the observed heterozygosity ranged from 0.208 to 0.894 and from 0.431 to 0.896 respectively. The experimental results show that the microsatellite sequences obtained by the enrichment of magnetic beads are of high quality and the efficiency of developing microsatellite markers is high. The experiment of 5 ’anchored PCR method is simple and convenient, and the obtained primers have higher genetic diversity index. The primers developed by the two methods can be used to study the genetic diversity of wild germplasm resources of Rehmannia.